This outstanding proposal plans to study and contrast the role of purinergic actions and cholinergic actions in the bladder. With evidence suggesting some gender specificity in purinergic functional significance, the project will also investigate mechanisms involved in sex hormone regulation of purinergic activity in the bladder. The majority of the work will be done in rats with some investigation performed on strips from human bladders. The methods to investigate these questions are in vivo cystometry in conscious, unrestrained rats, mRNA analysis of bladder tissue from rats, and ligand binding studies. In vivo studies include conscious animal studies to compare and contrast the relative importance of purinergic and cholinergic innervation of the bladder. In vitro whole bladder preparations for similar comparisons will complement the in vivo studies. In vitro studies of receptor subtypes using specific agonists, ecto-ATPase antagonists, and mRNA and ligand binding studies will provide information about the presence of receptor subtypes and the mechanisms involved in gender differences in purinergic responsiveness in the bladder. Calcium studies will be performed to determine the involvement of calcium in purinergic responses. Lastly, in vitro studies of rat and human bladder strip contractility will be done to compare animals and human tissues in males and females to determine any gender specificity.